Korean Journal of Clinical Laboratory Science (대한임상검사과학회지)

Korean Society for Clinical Laboratory Science (대한임상검사과학회)
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Mass Spectrometry-based Comparative Analysis of Membrane Protein: High-speed Centrifuge Method Versus Reagent-based Method

질량분석기를 활용한 막 단백질 비교분석: High-speed Centrifuge법과 Reagent-based법

  • Lee, Jiyeong (Department of Biomedical Laboratory Science, Eulji University) ;
  • Seok, Ae Eun (Department of Biomedical Laboratory Science, Eulji University) ;
  • Park, Arum (Department of Biomedical Laboratory Science, Eulji University) ;
  • Mun, Sora (Department of Senior Healthcare, BK21 Plus Program, Graduate School, Eulji University) ;
  • Kang, Hee-Gyoo (Department of Biomedical Laboratory Science, Eulji University)
  • 이지영 (을지대학교 보건과학대학 임상병리학과) ;
  • 석애은 (을지대학교 보건과학대학 임상병리학과) ;
  • 박아름 (을지대학교 보건과학대학 임상병리학과) ;
  • 문소라 (을지대학교 일반대학원 시니어헬스케어학과) ;
  • 강희규 (을지대학교 보건과학대학 임상병리학과)
  • Received : 2018.11.17
  • Accepted : 2019.02.18
  • Published : 2019.03.31
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Abstract

Membrane proteins are involved in many common diseases, including heart disease and cancer. In various disease states, such as cancer, abnormal signaling pathways that are related to the membrane proteins cause the cells to divide out of control and the expression of membrane proteins can be altered. Membrane proteins have the hydrophobic environment of a lipid bilayer, which makes an analysis of the membrane proteins notoriously difficult. Therefore, this study evaluated the efficacy of two different methods for optimal membrane protein extraction. High-speed centrifuge and reagent-based method with a -/+ filter aided sample preparation (FASP) were compared. As a result, the high-speed centrifuge method is quite effective in analyzing the mitochondrial inner membranes, while the reagent-based method is useful for endoplasmic reticulum membrane analysis. In addition, the function of the membrane proteins extracted from the two methods were analyzed using GeneGo software. GO processes showed that the endoplasmic reticulum-related responses had higher significance in the reagent-based method. An analysis of the process networks showed that one cluster in the high-speed centrifuge method and four clusters in the reagent-based method were visualized. In conclusion, the two methods are useful for the analysis of different subcellular membrane proteins, and are expected to assist in selecting the membrane protein extraction method by considering the target subcellular membrane proteins for study.

막 단백질은 심장질환, 암과 같은 우리 주변에서 흔히 발생하는 질병에 관련되어 있다. 이러한 암과 같은 특정한 질환 상태에서, 막 단백질과 관련된 신호 전달의 비정상은 세포분열을 통제하지 못하고 증가시킬 수 있으며 막 단백질의 발현에 변화가 생긴다. 막 단백질은 지질 이중층으로 이루어진 소수성 환경을 가지고 있어 불안정하기 때문에 막 단백질을 추출해서 연구를 수행하는데 어려움이 있다. 이번 연구에서는 최적화된 막 단백질 추출법을 확인하고자 서로 다른 두 가지 추출법의 효율성을 평가하였다. 두 가지 방법으로, high-speed centrifuge법과 reagent법이 비교되었다. 비교 분석결과, 미토콘드리아 내막 단백질 분석에는 high-speed centrifuge법이 효율적이고, 소포체 막 단백질 분석에는 reagent법이 유용함을 확인하였다. 게다가 유전자 온톨로지 소프트웨어를 이용해서 추출된 막 단백질의 기능분석을 진행하였을 때, 유전자 온톨로지는 reagent법에서 소포체 막 단백질에 연관된 반응이 활성화 되는 것을 확인할 수 있었다. 프로세스 네트워크 분석에서, high-speed centrifuge법에서는 하나의 클러스터를 형성화는 반면, reagent법에서는 네 개의 클러스터를 형성하는 것을 시각화하여 확인하였다. 결론적으로, 두 가지 분석법은 서로 다른 하위 막 단백질의 분석에 유용함을 확인할 수 있었다. 이러한 결과를 토대로, 막 단백질을 분석할 때, 표적의 세부 막 단백질을 고려하여 방법론을 선택하는데 도움을 줄 것으로 기대된다.

Keywords

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